The signals regulating human B cell functional differentiation have not been thoroughly explored. The research has focused on the identification of human B cell subsets and abnormalities in these subsets in human autoimmune disease. We have also examined the impact of signaling through B cell surface molecules, including CD40 and surface IgM/IgD, on B cell differentiation and the impact of regulators of these signaling pathways, such as vitamin D on B cell function. On particular interest has been the CD40 signaling cascade that regulates essential B cell maturational events such as germinal center formation, immunoglobulin heavy chain class switching, somatic hypermutation and differentiation to memory cells or plasma cells. One of the goals of the research has been to understand how CD40 signals and other signaling cascades impact human B cell differentiation so that one can understand abnormalities in these pathways that occur in autoimmune diseases such as systemic lupus erythematosus, rheumatoid arthritis and Sjogren's synfrome. Mechanisms connecting cell surface occupancy with downstream signaling events, specific gene transcription and functional outcomes are examined following engagement of surface receptors on B cells with recombinant ligands or monoclonal antibodies. Signaling cascades are dissected using novel biochemical and multi-parameter flow cytometric assays following transfection or transduction of B cells with constructs expressing constitutively active or dominant negative proteins. Gene expression is examined by microarray and confirmed by quantitative PCR, Western blotting, the proteomic technique Immunoaffinity Capillary Electrophoresis (ICE) and flow cytometry. Functional outcomes are examined in vitro, ex vivo immediately following isolation of human blood or tonsillar tissue samples, and following transplantation of activated human secondary lymphoid tissue and primary lymphocyte populations into the RAG KO/NOD/Perforin KO mouse model. Despite the complexity, specific checkpoints within the cascades regulate signaling and provide evidence of novel means to regulate B cell function therapeutically. In summary, the results of the experiments have delineated the role of CD40 signaling and other signaling events in abnormal germinal center reactions in patients with systemic lupus erythematosus and Sjogren's Syndrome that lead to differentiation of autoimmune plasma cell and memory B cell populations. Further delineation of the differentiation of human B cell subsets and abnormalities in human autoimmune disease should provide unique targets that could be exploited to regulate B cell activity in systemic lupus erythematosus, rheumatoid arthritis and Sjogren's syndrome..